The present invention relates to novel nucleic acid sequences encoding a member of the neuregulin family and to polypeptides encoded by the nucleic acid sequences.
The ErbB/HER family of growth factor receptors plays an important role in cell signaling during development and proliferation. Several ErbB/HER family members are associated with human diseases, including, for example, cancer. Numerous ligands which bind and activate ErbB/HER receptors have been identified, including epidermal growth factor (EGF), epiregulin, amphiregulin and members of the neuregulin family (Ben-Baruch et al., Hormones and growth factors in development and neoplasia. Dickson RB and Salomon D S (eds). Kulwer Academic Publishers: Boston, pp. 145-168, 1988; Burden and Yarden, Neuron 18:847-855). Because signal transduction pathways involving ErbB/HER family members are associated with human disease, there is a need in the art for ligands which signal through these receptors. The ligand proteins and the genes encoding them are useful for therapeutic purposes.
This invention provides a gene of the neuregulin family and polypeptides encoded by the gene. The invention is directed to mRNA expressed by mammalian cells, polynucleotides having coding regions corresponding to the mRNAs, protein and polypeptide products of the polynucleotides and mRNAs, and biological functions of the polypeptides and proteins.
It is an object of the invention to provide EGFH2 polynucleotides and polypeptides.
One embodiment of the invention provides a composition comprising an isolated nucleic acid molecule comprising a polynucleotide selected from the group consisting of:
(a) a polynucleotide encoding amino acids from about 1 to about 115 of SEQ ID NO:2;
(b) a polynucleotide encoding amino acids from about 2 to about 115 of SEQ ID NO:2;
(c) a polynucleotide encoding amino acids from about 1 to about 115 of SEQ ID NO:4;
(d) a polynucleotide encoding amino acids from about 2 to about 115 of SEQ ID NO:4;
(e) the polynucleotide complement of the polynucleotide of (a), (b), (c), or (d); and
(f) a polynucleotide at least 90% identical to the polynucleotide of (a), (b), (c), (d) or (e).
Another embodiment of the invention provides a composition comprising an isolated nucleic acid molecule comprising about 345 contiguous nucleotides from the coding region of SEQ ID NO:1 or SEQ ID NO:3.
In another embodiment, the invention provides a composition comprising an isolated nucleic acid molecule comprising a polynucleotide encoding a polypeptide wherein, except for at least one conservative amino acid substitution, said polypeptide has an amino acid sequence selected from the group consisting of:
(a) amino acids from about 1 to about 115 of SEQ ID NO:2;
(b) amino acids from about 2 to about 115 of SEQ ID NO:2;
(c) amino acids from about 1 to about 115 of SEQ ID NO:4;
(d) amino acids from about 2 to about 115 of SEQ ID NO:4.
In a preferred embodiment of the invention, the isolated nucleic acid molecule is DNA.
Another embodiment of the invention provides a method of making a recombinant vector comprising inserting the nucleic acid molecule into a vector in operable linkage to a promoter. The invention further provides a recombinant vector produced by this method.
In another embodiment, the invention provides a method of making a recombinant host cell comprising introducing recombinant vector into a host cell. The invention further provides a host cell produced by this method.
A further object of the invention is to provide a recombinant method of producing a polypeptide comprising culturing the recombinant host cell under conditions such that said polypeptide is expressed and recovering said polypeptide.
The present invention provides an isolated polypeptide comprising amino acids at least 95% identical to amino acids selected from the group consisting of:
(a) amino acids from about 1 to about 115 of SEQ ID NO:2;
(b) amino acids from about 2 to about 115 of SEQ ID NO:2;
(c) amino acids from about 1 to about 115 of SEQ ID NO:4; and
(d) amino acids from about 2 to about 115 of SEQ ID NO:4.
A preferred embodiment of the invention provides an isolated polypeptide wherein, except for at least one conservative amino acid substitution, said polypeptide has an amino acid sequence selected from the group consisting of (a), (b), (c), or (d) above.
In a most preferred embodiment, the invention provides an isolated polypeptide comprising amino acids selected from the group consisting of (a), (b), (c), or (d) above.
In a most preferred embodiment, the polypeptides comprise a biologically active EGFH2 polypeptide. In a preferred embodiment, the polypeptides comprise a precursor or mature EGFH2 protein. In another preferred embodiment, the polypeptides comprise an EGFH2 EGF-like domain.
A further embodiment of the invention provides a polypeptide comprising an epitope-bearing portion of a polypeptide selected from SEQ ID NO:2 or SEQ ID NO:4.
A preferred embodiment provides an epitope-bearing portion of the polypeptide, which comprises between about 10 and 100 contiguous amino acids of SEQ ID NO:2 or SEQ ID NO:4.
A more preferred embodiment of the invention provides an epitope-bearing portion of the polypeptide which comprises between about 12 and 50 contiguous amino acids of SEQ ID NO:2 or SEQ ID NO:4.
A most preferred embodiment of the invention provides an epitope-bearing portion of the polypeptide which comprises between about 15 and 25 contiguous amino acids of SEQ ID NO:2 or SEQ ID NO:4.
The invention further provides a composition comprising an isolated antibody that binds specifically to polypeptides of the present invention.
In preferred embodiments, the antibodies are monoclonal, polyclonal or single-chain antibodies.
An object of the invention is to provide a method for detecting EGFH2 polypeptides or mRNA for diagnostic and prognostic purposes.
In a preferred embodiment, the method comprises an antibody which binds a polynucleotide of the invention, contacting the antibody with a biological sample from a human suspected of having a EGFH2 protein-modulated disorder under binding conditions to form a duplex, and determining the amount of said duplex formed, compared to a normal sample.
In another preferred embodiment, the method comprises a polynucleotide that binds to mRNA encoding a polypeptide of the invention under stringent conditions, contacting nucleic acid of said sample with said polynucleotide under binding conditions to form a duplex, and determining the amount of said duplex formed, compared to a normal sample.
Another object of the invention is to provide methods of modulating the amount of EGFH2 protein in a subject, using the polypeptide and polynucleotide compositions of this invention.
In one embodiment, the method comprises administering an effective amount of a composition comprising the polypeptide of the invention or an antibody that binds to a polypeptide of the invention.
In another embodiment, the method comprises administering an effective amount of a composition consisting of a polynucleotide of the invention.
A further object of the invention is to provide methods for treating a EGFH2 protein-modulated disorder in a subject, using the polypeptide and polynucleotide compositions of this invention.
In one embodiment, the method comprises administering an effective amount of a composition comprising the polypeptide of the invention or an antibody that binds to a polypeptide of the invention, wherein said composition further comprises a pharmaceutically acceptable carrier.
In another embodiment, the method comprises administering an effective amount of a composition consisting of a polynucleotide of the invention, wherein said composition further comprises a pharmaceutically acceptable carrier.
In a preferred embodiment, the method is accomplished by implanting cells containing a polynucleotide expressing a polypeptide of the invention into the patient, wherein said cells express EGFH2 polypeptide in the patient.
In a most preferred embodiment, the implanted cells are encapsulated in a semipermeable membrane.
In another embodiment of the invention, patients are treated with a therapeutically effective amount of a polynucleotide capable of hybridizing to a polynucleotide of the invention or complement thereof.
In preferred embodiments, the polynucleotide is an antisense construct or a ribozyme.
In another preferred embodiment, the polynucleotide is a retroviral vector comprising a promoter and polynucleotides of the invention or complements thereof.
In yet another embodiment, patients are treated with a therapeutically effective amount of polypeptides capable of binding a polypeptide of the invention.
In a most preferred embodiment, the polypeptides are antibodies.
In another preferred embodiment, the polypeptides are wild-type or mutant receptors for EGFH2.
In another embodiment of the invention, patients are treated with polypeptides of the invention.